Reversible binding of the anticancer drug KXO1 (tirbanibulin) to the colchicine-binding site of ?-tubulin explains KXO1's low clinical toxicity
Reversible binding of the anticancer drug KXO1 (tirbanibulin) to the colchicine-binding site of ?-tubulin explains KXO1's low clinical toxicity
复制标题
抗癌药物 KXO1（tirbanibulin）与 β-微管蛋白的秋水仙碱结合位点的可逆结合解释了 KXO1 的低临床毒性
DOI：
10.1074/jbc.ra119.010732
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发表时间：
2019-11-29
影响因子：
4.8
通讯作者：
Chen, Lijuan
中科院分区：
文献类型：
--
作者：
Niu, Lu;Yang, Jianhong;Chen, Lijuan
KXO1(tirbanibuin或KX2？391)是一种SRC原癌基因非受体酪氨酸激酶(SRC)的非ATP竞争性抑制剂，正被临床研究用于治疗多种癌症和光化性角化病。最近，KXO1也被证明对微管蛋白有强烈的抑制作用。有趣的是，与传统的微管蛋白靶向药物不同，KXO1在临床前和临床研究中表现出低毒性，但其原因仍不清楚，KXO1结合部位和KXO1与微管蛋白相互作用的其他细节也是如此。在此，以细胞为基础的实验表明，KXO1在低纳米摩尔浓度下诱导微管蛋白解聚和G(2)/M期细胞周期停滞，与秋水仙碱类似，用作阳性对照。生化实验结果，包括N，N-亚乙基双(碘乙酰胺)竞争实验结果显示，KXO1结合在-微管蛋白上的秋水仙碱结合部位，进一步被微管蛋白？KXO1复合体的晶体结构所证实。决议。结晶学数据的高质量电子密度图使我们能够明确确定KXO1在秋水仙素结合部位的位置和方向，揭示KXO1与微管蛋白之间的详细相互作用。我们还发现KXO_1可逆地与纯化的微管蛋白结合，诱导完全可逆的细胞效应(G(2)/M期停滞)，并且在药物洗脱后5天没有细胞毒性，这说明KXO_1的S毒性很低。综上所述，我们发现KXO1结合到微管蛋白的秋水仙素结合部位，并分解了微管蛋白KXO1复合体的晶体结构。重要的是，KXO1的S可逆结合微管蛋白解释了它的临床低毒性，这一见解可以指导KXO1的进一步临床应用。
KXO1 (tirbanibulin or KX2?391) is as a non-ATP-competitive inhibitor of SRC proto-oncogene nonreceptor tyrosine kinase (SRC) and is being clinically investigated for the management of various cancers and actinic keratosis. Recently, KXO1 has also been shown to strongly inhibit tubulin. Interestingly, unlike conventional tubulin-targeting drugs, KXO1 has exhibited low toxicity in preclinical and clinical studies, but the reason for this remains elusive, as are the KXO1-binding site and other details of the interaction of KXO1 with tubulin. Here, cell-based experiments revealed that KXO1 induces tubulin depolymerization and G(2)/M phase cell cycle arrest at low nanomolar concentrations, similar to colchicine, used as a positive control. Results from biochemical experiments, including an N,N-ethylenebis(iodoacetamide) competition assay, disclosed that KXO1 binds to the colchicine-binding site on ?-tubulin, further confirmed by the crystal structure of the tubulin?KXO1 complex at 2.5-? resolution. A high-quality electron density map of the crystallographic data enabled us to unambiguously determine the position and orientation of KXO1 in the colchicine-binding site, revealing the detailed interactions between KXO1 and tubulin. We also found that KXO1 binds reversibly to purified tubulin, induces a totally reversible cellular effect (G(2)/M cell cycle arrest), and possesses no cellular toxicity 5 days after drug washout, explaining KXO1's low toxicity. In summary, we show that KXO1 binds to the colchicine-binding site of tubulin and resolved the crystal structure of the tubulin?KXO1 complex. Importantly, KXO1's reversible binding to tubulin explains its clinically low toxicity, an insight that could guide further clinical applications of KXO1.